An automated Raman-based platform for the sorting of live cells by functional properties
An automated Raman-based platform for the sorting of live cells by functional properties
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DOI：
10.1038/s41564-019-0394-9
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发表时间：
2019-06-01
影响因子：
28.3
通讯作者：
Stocker, Roman
中科院分区：
文献类型：
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作者：
Lee, Kang Soo;Palatinszky, Marton;Stocker, Roman
稳定同位素探测被广泛用于研究微生物类群在其自然环境中的功能，但从复杂样品中分选同位素标记的微生物细胞用于随后的基因组分析或培养仍处于早期阶段。在这里，我们介绍了一种用于自动分选稳定同位素探针标记的微生物细胞的光流体平台，该平台结合了微流体、光镊和拉曼显微光谱，产生了适合于随后的单细胞基因组学、微型宏基因组学或培养的活细胞。我们描述了这种拉曼激活的细胞分选方法的设计和优化，说明了它的操作与四个模型细菌（两个肠道，一个土壤和一个海洋），并证明其高分选精度（98.3 +/- 1.7%），吞吐量（200-500个细胞h（-1）; 3.3-8.3个细胞min（-1））和与培养的兼容性。应用这种分选方法对参与小鼠结肠中粘蛋白降解的细菌进行宏基因组表征，揭示了细菌的多样性，包括未充分研究的Muribaculaceae家族的几个成员，突出了这种生态位的复杂性和拉曼激活细胞分选用于识别靶向过程中关键参与者的潜力。
Stable-isotope probing is widely used to study the function of microbial taxa in their natural environment, but sorting of isotopically labelled microbial cells from complex samples for subsequent genomic analysis or cultivation is still in its early infancy. Here, we introduce an optofluidic platform for automated sorting of stable-isotope-probing-labelled microbial cells, combining microfluidics, optical tweezing and Raman microspectroscopy, which yields live cells suitable for subsequent single-cell genomics, mini-metagenomics or cultivation. We describe the design and optimization of this Raman-activated cell-sorting approach, illustrate its operation with four model bacteria (two intestinal, one soil and one marine) and demonstrate its high sorting accuracy (98.3 +/- 1.7%), throughput (200-500 cells h(-1); 3.3-8.3 cells min(-1)) and compatibility with cultivation. Application of this sorting approach for the metagenomic characterization of bacteria involved in mucin degradation in the mouse colon revealed a diverse consortium of bacteria, including several members of the underexplored family Muribaculaceae, highlighting both the complexity of this niche and the potential of Raman-activated cell sorting for identifying key players in targeted processes.